Transcription Analysis of Genes Encoding Homologues of Reductive Dehalogenases in “ Dehalococcoides ” sp. Strain CBDB1 by Using Terminal Restriction Fragment Length Polymorphism and Quantitative PCR
- 1 April 2009
- journal article
- Published by American Society for Microbiology in Applied and Environmental Microbiology
- Vol. 75 (7), 1876-1884
- https://doi.org/10.1128/aem.01042-08
Abstract
The transcription of reductive dehalogenase homologous ( rdh ) genes of “ Dehalococcoides ” sp. strain CBDB1 was investigated during the growth and reductive dechlorination of 1,2,3- and 1,2,4-trichlorobenzene (TCB). A method was developed to monitor the expression of all 32 rdhA genes present in the genome based on reverse transcription-PCR amplification with 13 degenerate primer pairs and terminal restriction fragment length polymorphism (t-RFLP) analysis. With this approach, the upregulation of the transcription of 29 rdhA genes was indicated in response to 1,2,3- and 1,2,4-TCB added after a substrate depletion period of 72 h. The transcription of the remaining three rdhA genes additionally was detected using specific primers. While most rdhA genes were upregulated similarly in cultures after induction with 1,2,3-TCB or 1,2,4-TCB, three rdhA genes responded differentially to 1,2,3- and 1,2,4-TCB, as revealed by the comparison of t-RFLP profiles. The enhanced transcription of cbdbA1453 and cbdbA187 was observed in the presence of 1,2,3-TCB, while the transcription of cbdbA1624 was strongly induced by 1,2,4-TCB. Comparison of t-RFLP profiles obtained from cDNA and genomic DNA indicated a particularly high induction of the transcription of cbrA (=cbdbA84) by both TCBs. As indicated by reverse transcription-quantitative PCR, the transcription of these plus two other rdhA genes (cbdbA1588 and cbdbA1618) increased within 48 to 72 h by one or two orders of magnitude. Subsequently, transcript levels slowly decreased and approached initial transcript levels several days after complete dehalogenation. Finally, cbrA was transcribed to a level of 22 transcripts per cbrA gene, suggesting that cbrA mRNA could be an appropriate biomarker for the investigation of the natural dechlorination potential at chlorobenzene-contaminated sites.Keywords
This publication has 45 references indexed in Scilit:
- Quantifying Genes and Transcripts To Assess the In Situ Physiology of “ Dehalococcoides ” spp. in a Trichloroethene-Contaminated Groundwater SiteApplied and Environmental Microbiology, 2008
- Identification of a Chlorobenzene Reductive Dehalogenase in Dehalococcoides sp. Strain CBDB1Applied and Environmental Microbiology, 2007
- Impact of Plant Functional Group, Plant Species, and Sampling Time on the Composition of nirK -Type Denitrifier Communities in SoilApplied and Environmental Microbiology, 2007
- Improved accuracy in terminal restriction fragment length polymorphism phylogenetic analysis using a novel internal size standard definitionOral Microbiology and Immunology, 2007
- Expression of Reductive Dehalogenase Genes in Dehalococcoides ethenogenes Strain 195 Growing on Tetrachloroethene, Trichloroethene, or 2,3-DichlorophenolApplied and Environmental Microbiology, 2007
- Comparative Proteomics of Dehalococcoides spp. Reveals Strain-Specific Peptides Associated with ActivityApplied and Environmental Microbiology, 2007
- Reductive Dehalogenase Gene Expression as a Biomarker for Physiological Activity of Dehalococcoides sppApplied and Environmental Microbiology, 2006
- Temporal Expression of Respiratory Genes in an Enrichment Culture Containing Dehalococcoides ethenogenesApplied and Environmental Microbiology, 2006
- Genetic Identification of a Putative Vinyl Chloride Reductase in Dehalococcoides sp. Strain BAV1Applied and Environmental Microbiology, 2004
- Molecular Identification of the Catabolic Vinyl Chloride Reductase from Dehalococcoides sp. Strain VS and Its Environmental DistributionApplied and Environmental Microbiology, 2004